Soybean cultivar S030159

ABSTRACT

A novel soybean cultivar, designated S030159, is disclosed. The invention relates to the seeds of soybean cultivar S030159, to the plants of soybean S030159 and to methods for producing a soybean plant produced by crossing the cultivar S030159 with itself or another soybean variety. The invention further relates to hybrid soybean seeds and plants produced by crossing the cultivar S030159 with another soybean cultivar.

BACKGROUND OF THE INVENTION

The present invention relates to a new and distinctive soybean cultivar,designated S030159. There are numerous steps in the development of anynovel, desirable plant germplasm. Plant breeding begins with theanalysis and definition of problems and weaknesses of the currentgermplasm, the establishment of program goals, and the definition ofspecific breeding objectives. The next step is selection of germplasmthat possess the traits to meet the program goals. The goal is tocombine in a single variety an improved combination of desirable traitsfrom the parental germplasm. These important traits may include higherseed yield, resistance to diseases and insects, better stems and roots,tolerance to drought and heat, and better agronomic quality.

Choice of breeding or selection methods depends on the mode of plantreproduction, the heritability of the trait(s) being improved, and thetype of cultivar used commercially (e.g., F₁ hybrid cultivar, purelinecultivar, etc.). For highly heritable traits, a choice of superiorindividual plants evaluated at a single location will be effective,whereas for traits with low heritability, selection should be based onmean values obtained from replicated evaluations of families of relatedplants. Popular selection methods commonly include pedigree selection,modified pedigree selection, mass selection, and recurrent selection.

The complexity of inheritance influences choice of the breeding method.Backcross breeding is used to transfer one or a few favorable genes fora highly heritable trait into a desirable cultivar. This approach hasbeen used extensively for breeding disease-resistant cultivars. Variousrecurrent selection techniques are used to improve quantitativelyinherited traits controlled by numerous genes. The use of recurrentselection in self-pollinating crops depends on the ease of pollination,the frequency of successful hybrids from each pollination, and thenumber of hybrid offspring from each successful cross.

Each breeding program should include a periodic, objective evaluation ofthe efficiency of the breeding procedure. Evaluation criteria varydepending on the goal and objectives, but should include gain fromselection per year based on comparisons to an appropriate standard,overall value of the advanced breeding lines, and number of successfulcultivars produced per unit of input (e.g., per year, per dollarexpended, etc.).

Promising advanced breeding lines are thoroughly tested and compared toappropriate standards in environments representative of the commercialtarget area(s) for three or more years. The best lines are candidatesfor new commercial cultivars; those still deficient in a few traits maybe used as parents to produce new populations for further selection.

These processes, which lead to the final step of marketing anddistribution, usually take from eight to 12 years from the time thefirst cross is made. Therefore, development of new cultivars is atime-consuming process that requires precise forward planning, efficientuse of resources, and a minimum of changes in direction.

A most difficult task is the identification of individuals that aregenetically superior, because for most traits the true genotypic valueis masked by other confounding plant traits or environmental factors.One method of identifying a superior plant is to observe its performancerelative to other experimental plants and to a widely grown standardcultivar. If a single observation is inconclusive, replicatedobservations provide a better estimate of its genetic worth.

The goal of plant breeding is to develop new, unique and superiorsoybean cultivars and hybrids. The breeder initially selects and crossestwo or more parental lines, followed by repeated selfing and selection,producing many new genetic combinations. The breeder can theoreticallygenerate billions of different genetic combinations via crossing,selfing and mutations. The breeder has no direct control at the cellularlevel. Therefore, two breeders will never develop the same line, or evenvery similar lines, having the same soybean traits.

Each year, the plant breeder selects the germplasm to advance to thenext generation. This germplasm is grown under unique and differentgeographical, climatic and soil conditions, and further selections arethen made, during and at the end of the growing season. The cultivarswhich are developed are unpredictable. This unpredictability is becausethe breeder's selection occurs in unique environments, with no controlat the DNA level (using conventional breeding procedures), and withmillions of different possible genetic combinations being generated. Abreeder of ordinary skill in the art cannot predict the final resultinglines he develops, except possibly in a very gross and general fashion.The same breeder cannot produce the same cultivar twice by using theexact same original parents and the same selection techniques. Thisunpredictability results in the expenditure of large amounts of researchmonies to develop superior new soybean cultivars.

The development of new soybean cultivars requires the development andselection of soybean varieties, the crossing of these varieties andselection of superior hybrid crosses. The hybrid seed is produced bymanual crosses between selected male-fertile parents or by using malesterility systems. These hybrids are selected for certain single genetraits such as pod color, flower color, pubescence color or herbicideresistance which indicate that the seed is truly a hybrid. Additionaldata on parental lines, as well as the phenotype of the hybrid,influence the breeder's decision whether to continue with the specifichybrid cross.

Pedigree breeding and recurrent selection breeding methods are used todevelop cultivars from breeding populations. Breeding programs combinedesirable traits from two or more cultivars or various broad-basedsources into breeding pools from which cultivars are developed byselfing and selection of desired phenotypes. The new cultivars areevaluated to determine which have commercial potential.

Pedigree breeding is used commonly for the improvement ofself-pollinating crops. Two parents which possess favorable,complementary traits are crossed to produce an F₁. An F₂ population isproduced by selfing one or several F₁'s. Selection of the bestindividuals may begin in the F₂ population; then, beginning in the F₃,the best individuals in the best families are selected. Replicatedtesting of families can begin in the F₄ generation to improve theeffectiveness of selection for traits with low heritability. At anadvanced stage of inbreeding (i.e., F₆ and F₇), the best lines ormixtures of phenotypically similar lines are tested for potentialrelease as new cultivars.

Mass and recurrent selections can be used to improve populations ofeither self- or cross-pollinating crops. A genetically variablepopulation of heterozygous individuals is either identified or createdby intercrossing several different parents. The best plants are selectedbased on individual superiority, outstanding progeny, or excellentcombining ability. The selected plants are intercrossed to produce a newpopulation in which further cycles of selection are continued.

Backcross breeding has been used to transfer genes for a simplyinherited, highly heritable trait into a desirable homozygous cultivaror inbred line which is the recurrent parent. The source of the trait tobe transferred is called the donor parent. The resulting plant isexpected to have the attributes of the recurrent parent (e.g., cultivar)and the desirable trait transferred from the donor parent. After theinitial cross, individuals possessing the phenotype of the donor parentare selected and repeatedly crossed (backcrossed) to the recurrentparent. The resulting plant is expected to have the attributes of therecurrent parent (e.g., cultivar) and the desirable trait transferredfrom the donor parent.

The single-seed descent procedure in the strict sense refers to plantinga segregating population, harvesting a sample of one seed per plant, andusing the one-seed sample to plant the next generation. When thepopulation has been advanced from the F₂ to the desired level ofinbreeding, the plants from which lines are derived will each trace todifferent F₂ individuals. The number of plants in a population declineseach generation due to failure of some seeds to germinate or some plantsto produce at least one seed. As a result, not all of the F₂ plantsoriginally sampled in the population will be represented by a progenywhen generation advance is completed.

In a multiple-seed procedure, soybean breeders commonly harvest one ormore pods from each plant in a population and thresh them together toform a bulk. Part of the bulk is used to plant the next generation andpart is put in reserve. The procedure has been referred to as modifiedsingle-seed descent or the pod-bulk technique.

The multiple-seed procedure has been used to save labor at harvest. Itis considerably faster to thresh pods with a machine than to remove oneseed from each by hand for the single-seed procedure. The multiple-seedprocedure also makes it possible to plant the same number of seeds of apopulation each generation of inbreeding. Enough seeds are harvested tomake up for those plants that did not germinate or produce seed.

Descriptions of other breeding methods that are commonly used fordifferent traits and crops can be found in one of several referencebooks (e.g., Allard, 1960; Simmonds, 1979; Sneep et al., 1979; Fehr,1987).

Proper testing should detect any major faults and establish the level ofsuperiority or improvement over current cultivars. In addition toshowing superior performance, there must be a demand for a new cultivarthat is compatible with industry standards or which creates a newmarket. The introduction of a new cultivar will incur additional coststo the seed producer, the grower, processor and consumer; for specialadvertising and marketing, altered seed and commercial productionpractices, and new product utilization. The testing preceding release ofa new cultivar should take into consideration research and developmentcosts as well as technical superiority of the final cultivar. Forseed-propagated cultivars, it must be feasible to produce seed easilyand economically.

Soybean, Glycine max (L), is an important and valuable field crop. Thus,a continuing goal of plant breeders is to develop stable, high yieldingsoybean cultivars that are agronomically sound. The reasons for thisgoal are obviously to maximize the amount of grain produced on the landused and to supply food for both animals and humans. To accomplish thisgoal, the soybean breeder must select and develop soybean plants thathave the traits that result in superior cultivars.

SUMMARY OF THE INVENTION

According to the invention, there is provided a novel soybean cultivar,designated S030159. This invention thus relates to the seeds of soybeancultivar S030159, to the plants of soybean S030159 and to methods forproducing a soybean plant produced by crossing the soybean S030159 withitself or another soybean line, and the creation of variants bymutagenesis or transformation of soybean S030159.

Thus, any such methods using the soybean variety S030159 are part ofthis invention: selfing, backcrosses, hybrid production, crosses topopulations, and the like. All plants produced using soybean varietyS030159 as a parent are within the scope of this invention.Advantageously, the soybean variety could be used in crosses with other,different, soybean plants to produce first generation (F₁) soybeanhybrid seeds and plants with superior characteristics.

In another aspect, the present invention provides for single or multiplegene converted plants of S030159. The transferred gene(s) may preferablybe a dominant or recessive allele. Preferably, the transferred gene(s)will confer such traits as herbicide resistance, insect resistance,resistance for bacterial, fungal, or viral disease, male fertility, malesterility, enhanced nutritional quality, and industrial usage. The genemay be a naturally occurring soybean gene or a transgene introducedthrough genetic engineering techniques.

In another aspect, the present invention provides regenerable cells foruse in tissue culture of soybean plant S030159. The tissue culture willpreferably be capable of regenerating plants having the physiologicaland morphological characteristics of the foregoing soybean plant, and ofregenerating plants having substantially the same genotype as theforegoing soybean plant. Preferably, the regenerable cells in suchtissue cultures will be embryos, protoplasts, meristematic cells,callus, pollen, leaves, anthers, roots, root tips, flowers, seeds, podsor stems. Still further, the present invention provides soybean plantsregenerated from the tissue cultures of the invention.

Definitions

In the description and tables which follow, a number of terms are used.In order to provide a clear and consistent understanding of thespecification and claims, including the scope to be given such terms,the following definitions are provided:

-   -   Allele. Allele is any of one or more alternative forms of a        gene, all of which alleles relate to one trait or        characteristic. In a diploid cell or organism, the two alleles        of a given gene occupy corresponding loci on a pair of        homologous chromosomes.    -   Backcrossing. Backcrossing is a process in which a breeder        repeatedly crosses hybrid progeny back to one of the parents,        for example, a first generation hybrid F₁ with one of the        parental genotypes of the F₁ hybrid.    -   Brown Stem Rot. This is a visual disease score from 1 to 9        comparing all genotypes in a given test. The score is based on        leaf symptoms of yellowing and necrosis caused by brown stem        rot. A score of 9 indicates no symptoms. Visual scores range to        a score of 1 which indicates severe symptoms of leaf yellowing        and necrosis.    -   Cotyledon. A cotyledon is a type of seed leaf. The cotyledon        contains the food storage tissues of the seed.    -   Embryo. The embryo is the small plant contained within a mature        seed.    -   Emergence. This score indicates the ability of the seed to        emerge when planted 3″ deep in sand and with a controlled        temperature of 25° C. The number of plants that emerge each day        are counted. Based on this data, each genotype is given a 1 to 9        score based on its rate of emergence and percent of emergence. A        score of 9 indicates an excellent rate and percent of emergence,        an intermediate score of 5 indicates average ratings and a 1        score indicates a very poor rate and percent of emergence.    -   Hilum. This refers to the scar left on the seed which marks the        place where the seed was attached to the pod prior to the seed        being harvested.    -   Hypocotyl. A hypocotyl is the portion of an embryo or seedling        between the cotyledons and the root. Therefore, it can be        considered a transition zone between shoot and root.    -   Iron-Deficiency Chlorosis. Plants are scored 1 to 9 based on        visual observations. A score of 9 means no stunting of the        plants or yellowing of the leaves and a score of 1 indicates the        plants are dead or dying caused by iron-deficiency chlorosis, a        score of 5 means plants have intermediate health with some leaf        yellowing.    -   Lodging Resistance. Lodging is rated on a scale of 1 to 9. A        score of 9 indicates erect plants. A score of 5 indicates plants        are leaning at a 45° angle in relation to the ground and a score        of 1 indicates plants are laying on the ground.    -   Maturity Date. Plants are considered mature when 95% of the pods        have reached their mature color. The number of days are either        calculated from August 31 or from the planting date.    -   Maturity Group. This refers to an agreed-on industry division of        groups of varieties, based on zones in which they are adapted        primarily according to day length or latitude. They consist of        very long day length varieties (Groups 000, 00, 0), and extend        to very short day length varieties (Groups VII, VIII, IX, X).    -   Relative Maturity (RM). The term relative maturity is a        numerical value that is assigned to a soybean variety based on        comparisons with the maturity values of other varieties. The        number preceding the decimal point in the RM refers to the        maturity group. The number following the decimal point refers to        the relative earliness or lateness within each maturity group.        For example, a 3.0 is an early group III variety, while a 3.9 is        a late group III variety.    -   Oil or oil percent. Soybean seeds contain a considerable amount        of oil. Oil is measured by NIR spectrophotometry, and is        reported on an as is percentage basis.    -   Oleic Acid Percent. Oleic acid is one of the five most abundant        fatty acids in soybean seeds. It is measured by gas        chromatography and is reported as a percent of the total oil        content.    -   Palmitic Acid Percent. Palmitic acid is one of the five most        abundant fatty acids in soybean seeds. It is measured by gas        chromatography and is reported as a percent of the total oil        content.    -   Phytophthora Tolerance. Tolerance to Phytophthora root rot is        rated on a scale of 1 to 9, with a score of 9 being the best or        highest tolerance ranging down to a score of 1 which indicates        the plants have no tolerance to Phytophthora.    -   Phenotlyic Score. The Phenotypic Score is a visual rating of        general appearance of the variety. All visual traits are        considered in the score including healthiness, standability,        appearance and freedom of disease. Ratings are scored from 1        being poor to 9 being excellent.    -   Plant Height. Plant height is taken from the top of soil to top        node of the plant and is measured in centimeters.    -   Pod. This refers to the fruit of a soybean plant. It consists of        the hull or shell (pericarp) and the soybean seeds.    -   Protein Percent. Soybean seeds contain a considerable amount of        protein. Protein is generally measured by NIR spectrophotometry,        and is reported on an as is percentage basis.    -   Pubescence. This refers to a covering of very fine hairs closely        arranged on the leaves, stems and pods of the soybean plant.    -   Quantitative Trait Loci (QTL). Quantitative trait loci (QTL)        refer to genetic loci that control to some degree numerically        representable traits that are usually continuously distributed.    -   Regeneration. Regeneration refers to the development of a plant        from tissue culture.    -   Seed Protein Peroxidase Activity. Seed protein peroxidase        activity is defined as a chemical taxonomic technique to        separate cultivars based on the presence or absence of the        peroxidase enzyme in the seed coat. There are two types of        soybean cultivars, those having high peroxidase activity (dark        red color) and those having low peroxidase activity (no color).    -   Seed Yield (Bushels/Acre). The yield in bushels/acre is the        actual yield of the grain at harvest.    -   Seeds per Pound. Soybean seeds vary in seed size, therefore, the        number of seeds required to make up one pound also varies. This        affects the pounds of seed required to plant a given area, and        can also impact end uses.    -   Shattering. The amount of pod dehiscence prior to harvest. Pod        dehiscence involves seeds falling from the pods to the soil.        This is a visual score from 1 to 9 comparing all genotypes        within a given test. A score of 9 means pods have not opened and        no seeds have fallen out. A score of 5 indicates approximately        50% of the pods have opened, with seeds falling to the ground        and a score of 1 indicates 100% of the pods are opened.    -   Single Gene Converted (Conversion). Single gene converted        (conversion) plant refers to plants which are developed by a        plant breeding technique called backcrossing wherein essentially        all of the desired morphological and physiological        characteristics of a variety are recovered in addition to the        single gene transferred into the variety via the backcrossing        technique or via genetic engineering.

DETAILED DESCRIPTION OF THE INVENTION

S030159 is a late maturity group II variety with resistance to Roundup™herbicide conferring tolerance to glyphosate herbicides. S030159 hasvery high yield potential when compared to lines of similar maturity andhas excellent agronomic characteristics including lodging resistance.

Some of the criteria used to select in various generations include: seedyield, lodging resistance, emergence, disease tolerance, maturity, lateseason plant intactness, plant height and shattering resistance.

The cultivar has shown uniformity and stability, as described in thefollowing variety description information. It has been self-pollinated asufficient number of generations with careful attention to uniformity ofplant type. The line has been increased with continued observation foruniformity.

Soybean cultivar S030159 has the following morphologic and othercharacteristics (based primarily on data collected at Adel, Iowa).

TABLE 1 VARIETY DESCRIPTION INFORMATION Seed Coat Color: Yellow SeedCoat Luster (Matured Hand Shelled Seed): Shiny Hilum Color: (MatureSeed) - Brown Cotyledon Color (Mature Seed): Yellow Leaflet Shape: OvateFlower Color: Purple Pod color: Tan Plant Pubescence Color: Light TawnyGrowth Habit: Indeterminate Maturity Group: II Relative Maturity: 2.8Plant Lodging Score: 6 Plant Height: 89 cm Seed Content: Protein: 35.2%;Oil: 19.5% Seed Size (# seeds/lb.): 3630 Physiological Responses:Roundup Ready ™ Herbicide: Resistant

Disease Resistance: Soybean Cyst Nematode—rhg1

This invention is also directed to methods for producing a soybean plantby crossing a first parent soybean plant with a second parent soybeanplant, wherein the first or second soybean plant is the soybean plantfrom the line S030159. Further, both first and second parent soybeanplants may be from the cultivar S030159. Therefore, any methods usingthe cultivar S030159 are part of this invention: selfing, backcrosses,hybrid breeding, and crosses to populations. Any plants produced usingcultivar S030159 as a parent are within the scope of this invention.

Useful methods include but are not limited to expression vectorsintroduced into plant tissues using a direct gene transfer method suchas microprojectile-mediated delivery, DNA injection, electroporation andthe like. More preferably expression vectors are introduced into planttissues using the microprojectile media delivery with the biolisticdevice Agrobacterium-medicated transformation. Transformant plantsobtained with the protoplasm of the invention are intended to be withinthe scope of this invention.

The cultivar S030159 is similar to DKB26-52. While similar to DKB26-52,there are numerous differences including: S030159 has brown hila whileDKB26-52 has imperfect black hila.

Further Embodiments of the Invention

With the advent of molecular biological techniques that have allowed theisolation and characterization of genes that encode specific proteinproducts, scientists in the field of plant biology developed a stronginterest in engineering the genome of plants to contain and expressforeign genes, or additional, or modified versions of native, orendogenous, genes (perhaps driven by different promoters) in order toalter the traits of a plant in a specific manner. Such foreignadditional and/or modified genes are referred to herein collectively as“transgenes”. Over the last fifteen to twenty years several methods forproducing transgenic plants have been developed, and the presentinvention, in particular embodiments, also relates to transformedversions of the claimed variety or line.

Plant transformation involves the construction of an expression vectorwhich will function in plant cells. Such a vector comprises DNAcomprising a gene under control of or operatively linked to a regulatoryelement (for example, a promoter). The expression vector may contain oneor more such operably linked gene/regulatory element combinations. Thevector(s) may be in the form of a plasmid, and can be used alone or incombination with other plasmids, to provide transformed soybean plants,using transformation methods as described below to incorporatetransgenes into the genetic material of the soybean plant(s).

Expression Vectors for Soybean Transformation: Marker Genes—Expressionvectors include at least one genetic marker, operably linked to aregulatory element (a promoter, for example) that allows transformedcells containing the marker to be either recovered by negativeselection, i.e., inhibiting growth of cells that do not contain theselectable marker gene, or by positive selection, i.e., screening forthe product encoded by the genetic marker. Many commonly used selectablemarker genes for plant transformation are well known in thetransformation arts, and include, for example, genes that code forenzymes that metabolically detoxify a selective chemical agent which maybe an antibiotic or a herbicide, or genes that encode an altered targetwhich is insensitive to the inhibitor. A few positive selection methodsare also known in the art.

One commonly used selectable marker gene for plant transformation is theneomycin phosphotransferase II (nptII) under the control of plantregulatory signals confers resistance to kanamycin. Fraley et al., Proc.Natl. Acad. Sci. U.S.A., 80:4803 (1983). Another commonly usedselectable marker gene is the hygromycin phosphotransferase gene whichconfers resistance to the antibiotic hygromycin. Vanden Elzen et al.,Plant Mol. Biol., 5:299 (1985).

Additional selectable marker genes of bacterial origin that conferresistance to antibiotics include gentamycin acetyl transferase,streptomycin phosphotransferase, aminoglycoside-3′-adenyl transferase,the bleomycin resistance determinant. Hayford et al., Plant Physiol.86:1216 (1988), Jones et al., Mol. Gen. Genet., 210:86 (1987), Svab etal., Plant Mol. Biol. 14:197 (1990). Hille et al., Plant Mol. Biol.7:171 (1986). Other selectable marker genes confer resistance toherbicides such as glyphosate, glufosinate or broxynil. Comai et al.,Nature 317:741-744 (1985), Gordon-Kamm et al., Plant Cell 2:603-618(1990) and Stalker et al., Science 242:419423 (1988).

Other selectable marker genes for plant transformation are not ofbacterial origin. These genes include, for example, mouse dihydrofolatereductase, plant 5-enolpyruvylshikimate-3-phosphate synthase and plantacetolactate synthase. Eichholtz et al., Somatic Cell Mol. Genet. 13:67(1987), Shah et al., Science 233:478 (1986), Charest et al., Plant CellRep. 8:643 (1990).

Another class of marker genes for plant transformation require screeningof presumptively transformed plant cells rather than direct geneticselection of transformed cells for resistance to a toxic substance suchas an antibiotic. These genes are particularly useful to quantify orvisualize the spatial pattern of expression of a gene in specifictissues and are frequently referred to as reporter genes because theycan be fused to a gene or gene regulatory sequence for the investigationof gene expression. Commonly used genes for screening presumptivelytransformed cells include β-glucuronidase (GUS, β-galactosidase,luciferase and chloramphenicol, acetyltransferase. Jefferson, R. A.,Plant Mol. Biol. Rep. 5:387 (1987), Teeri et al., EMBO J. 8:343 (1989),Koncz et al., Proc. Natl. Acad. Sci U.S.A. 84:131 (1987), DeBlock etal., EMBO J. 3:1681 (1984).

Recently, in vivo methods for visualizing GUS activity that do notrequire destruction of plant tissue have been made available. MolecularProbes publication 2908, Imagene Green™, p. 1-4 (1993) and Naleway etal., J. Cell Biol. 115:151a (1991). However, these in vivo methods forvisualizing GUS activity have not proven useful for recovery oftransformed cells because of low sensitivity, high fluorescentbackgrounds and limitations associated with the use of luciferase genesas selectable markers.

More recently, a gene encoding Green Fluorescent Protein (GFP) has beenutilized as a marker for gene expression in prokaryotic and eukaryoticcells. Chalfie et al., Science 263:802 (1994). GFP and mutants of GFPmay be used as screenable markers.

Promoters—Genes included in expression vectors must be driven bynucleotide sequence comprising a regulatory element, for example, apromoter. Several types of promoters are now well known in thetransformation arts, as are other regulatory elements that can be usedalone or in combination with promoters.

As used herein, “promoter” includes reference to a region of DNAupstream from the start of transcription and involved in recognition andbinding of RNA polymerase and other proteins to initiate transcription.A “plant promoter” is a promoter capable of initiating transcription inplant cells. Examples of promoters under developmental control includepromoters that preferentially initiate transcription in certain tissues,such as leaves, roots, seeds, fibers, xylem vessels, tracheids, orsclerenchyma. Such promoters are referred to as “tissue-preferred”.Promoters which intitiate transcription only in certain tissue arereferred to as “tissue-specific”. A “cell type” specific promoterprimarily drives expression in certain cell types in one or more organs,for example, vascular cells in roots or leaves. An “inducible” promoteris a promoter which is under environmental control. Examples ofenvironmental conditions that may effect transcription by induciblepromoters include anaerobic conditions or the presence of light.Tissue-specific, tissue-preferred, cell type specific, and induciblepromoters constitute the class of “non-constitutive” promoters. A“constitutive” promoter is a promoter which is active under mostenvironmental conditions.

A. Inducible Promoters—An inducible promoter is operably linked to agene for expression in soybean. Optionally, the inducible promoter isoperably linked to a nucleotide sequence encoding a signal sequencewhich is operably linked to a gene for expression in soybean. With aninducible promoter the rate of transcription increases in response to aninducing agent.

Any inducible promoter can be used in the instant invention. See Ward etal., Plant Mol. Biol. 22:361-366 (1993). Exemplary inducible promotersinclude, but are not limited to, that from the ACEI system whichresponds to copper (Mett et al., PNAS 90:45674571 (1993)); In2 gene frommaize which responds to benzenesulfonamide herbicide safeners (Hersheyet al., Mol. Gen Genetics 227:229-237 (1991) and Gatz et al., Mol. Gen.Genetics 243:32-38 (1994)) or Tet repressor from Tn10 (Gatz et al., Mol.Gen. Genetics 227:229-237 (1991). A particularly preferred induciblepromoter is a promoter that responds to an inducing agent to whichplants do not normally respond. An exemplary inducible promoter is theinducible promoter from a steroid hormone gene, the transcriptionalactivity of which is induced by a glucocorticosteroid hormone. Schenaiet al., Proc. Natl. Acad. Sci. U.S.A. 88:0421 (1991).

B. Constitutive Promoters—A constitutive promoter is operably linked toa gene for expression in soybean or the constitutive promoter isoperably linked to a nucleotide sequence encoding a signal sequencewhich is operably linked to a gene for expression in soybean.

Many different constitutive promoters can be utilized in the instantinvention. Exemplary constitutive promoters include, but are not limitedto, the promoters from plant viruses such as the 35S promoter from CaMV(Odell et al., Nature 313:810-812 (1985) and the promoters from suchgenes as rice actin (McElroy et al., Plant Cell(2:163-171 (1990));ubiquitin (Christensen et al., Plant Mol. Biol. 12:619-632 (1989) andChristensen et al., Plant Mol. Biol. 18:675-689 (1992)); pEMU (Last etal., Theor. Appl. Genet. 81:581-588 (1991)); MAS (Velten et al., EMBO J.3:2723-2730 (1984)) and maize H3 histone (Lepetit et al., Mol. Gen.Genetics 231:276-285 (1992) and Atanassova et al., Plant Journal 2 (3):291-300 (1992)).

The ALS promoter, Xba1/Ncol fragment 5′ to the Brassica napus ALS3structural gene (or a nucleotide sequence similarity to said Xba1/Ncolfragment), represents a particularly useful constitutive promoter. SeePCT application WO96/30530.

C. Tissue-specific or Tissue-preferred Promoters—A tissue-specificpromoter is operably linked to a gene for expression in soybean.Optionally, the tissue-specific promoter is operably linked to anucleotide sequence encoding a signal sequence which is operably linkedto a gene for expression in soybean. Plants transformed with a gene ofinterest operably linked to a tissue-specific promoter produce theprotein product of the transgene exclusively, or preferentially, in aspecific tissue.

Any tissue-specific or tissue-preferred promoter can be utilized in theinstant invention. Exemplary tissue-specific or tissue-preferredpromoters include, but are not limited to, a root-preferredpromoter—such as that from the phaseolin gene (Murai et al., Science23:476482 (1983) and Sengupta-Gopalan et al., Proc. Natl. Acad. Sci.U.S.A. 82:3320-3324 (1985)); a leaf-specific and light-induced promotersuch as that from cab or rubisco (Simpson et al., EMBO J.4(11):2723-2729 (1985) and Timko et al., Nature 318:579-582 (1985)); ananther-specific promoter such as that from LAT52 (Twell et al., Mol.Gen. Genetics 217:240-245 (1989)); a pollen-specific promoter such asthat from Zm 13 (Guerrero et al., Mol. Gen. Genetics 244:161-168 (1993))or a microspore-preferred promoter such as that from apg (Twell et al.,Sex. Plant Reprod. 6:217-224 (1993). Signal Sequences for TargetingProteins to Subcellular Compartments

Transport of protein produced by transgenes to a subcellular compartmentsuch as the chloroplast, vacuole, peroxisome, glyoxysome, cell wall ormitochondroin or for secretion into the apoplast, is accomplished bymeans of operably linking the nucleotide sequence encoding a signalsequence to the 5′ and/or 3′ region of a gene encoding the protein ofinterest. Targeting sequences at the 5′ and/or 3′ end of the structuralgene may determine, during protein synthesis and processing, where theencoded protein is ultimately compartmentalized.

The presence of a signal sequence directs a polypeptide to either anintracellular organelle or subcellular compartment or for secretion tothe apoplast. Many signal sequences are known in the art. See, forexample Becker et al., Plant Mol. Biol. 20:49 (1992), Close, P. S.,Master's Thesis, Iowa State University (1993), Knox, C., et al.,“Structure and Organization of Two Divergent Alpha-Amylase Genes fromBarley”, Plant Mol. Biol. 9:3-17 (1987), Lemeret al., Plant Physiol.91:124-129 (1989), Fontes et al., Plant Cell 3:483496 (1991), Matsuokaet al., Proc. Natl. Acad. Sci. 88:834 (1991), Gould et al., J. Cell.Biol. 108:1657 (1989), Creissen et al., Plant J. 2:129 (1991), Kalderon,et al., A short amino acid sequence able to specify nuclear location,Cell39:499-509 (1984), Steifel, et al., Expression of a maize cell wallhydroxyproline-rich glycoprotein gene in early leaf and root vasculardifferentiation, Plant Cell 2:785-793 (1990).

Foreign Protein Genes and Agronomic Genes—With transgenic plantsaccording to the present invention, a foreign protein can be produced incommercial quantities. Thus, techniques for the selection andpropagation of transformed plants, which are well understood in the art,yield a plurality of transgenic plants which are harvested in aconventional manner, and a foreign protein then can be extracted from atissue of interest or from total biomass. Protein extraction from plantbiomass can be accomplished by known methods which are discussed, forexample, by Heney and Orr, Anal. Biochem. 114:92-6 (1981).

According to a preferred embodiment, the transgenic plant provided forcommercial production of foreign protein is a soybean plant. In anotherpreferred embodiment, the biomass of interest is seed. For therelatively small number of transgenic plants that show higher levels ofexpression, a genetic map can be generated, primarily via conventionalRFLP, PCR and SSR analysis, which identifies the approximate chromosomallocation of the integrated DNA molecule. For exemplary methodologies inthis regard, see Glick and Thompson, Methods in Plant Molecular Biologyand Biotechnology CRC Press, Boca Raton 269:284 (1993). Map informationconcerning chromosomal location is useful for proprietary protection ofa subject transgenic plant. If unauthorized propagation is undertakenand crosses made with other germplasm, the map of the integration regioncan be compared to similar maps for suspect plants, to determine if thelatter have a common parentage with the subject plant. Map comparisonswould involve hybridizations, RFLP, PCR, SSR and sequencing, all ofwhich are conventional techniques.

Likewise, by means of the present invention, agronomic genes can beexpressed in transformed plants. More particularly, plants can begenetically engineered to express various phenotypes of agronomicinterest. Exemplary genes implicated in this regard include, but are notlimited to, those categorized below:

-   -   1. Genes That Confer Resistance to Pests or Disease and That        Encode:    -   A. Plant disease resistance genes. Plant defenses are often        activated by specific interaction between the product of a        disease resistance gene (R) in the plant and the product of a        corresponding avirulence (Avr) gene in the pathogen. A plant        variety can be transformed with cloned resistance gene to        engineer plants that are resistant to specific pathogen strains.        See, for example Jones et al., Science 266:789 (1994) (cloning        of the tomato Cf-9 gene for resistance to Cladosporium fulvum);        Martin et al., Science 262:1432 (1993) (tomato Pto gene for        resistance to Pseudomonas syringae pv. Tomato encoddes a protein        kinase); Mindrinos et al., Cell 78:1089 (1994) (Arabidopsis RSP2        gene for resistance to Pseudomonas syringae).    -   B. A gene conferring resistance to a pest, such as soybean cyst        nematode. See e.g., PCT Application WO96/30517; PCT Application        WO93/19181.    -   C. A Bacillus thuringiensis protein, a derivative thereof or a        synthetic polypeptide modeled thereon. See, for example, Geiser        et al., Gene 48:109 (1986), who disclose the cloning and        nucleotide sequence of a Bt δ-endotoxin gene. Moreover, DNA        molecules encoding δendotoxin genes can be purchased from        American Type Culture Collection, Manassas, Va., for example,        under ATCC Accession Nos. 40098, 67136, 31995 and 31998.    -   D. A lectin. See, for example, the disclose by Van Damme et al.,        Plant Molec. Biol. 24:25 (1994), who disclose the nucleotide        sequences of several Clivia miniata mannose-binding lectin        genes.    -   E. A vitamin-binding protein such as avidin. See PCT application        US93106487. The application teaches the use of avidin and avidin        homologues as larvicides against insect pests.    -   F. An enzyme inhibitor, for example, a protease or proteinase        inhibitor or an amylase inhibitor. See, for example, Abe et        al., J. Biol. Chem. 262:16793 (1987) (nucleotide sequence of        rice cysteine proteinase inhibitor), Huub et al., Plant Molec.        Biol. 21:985 (1993) (nucleotide sequence of cDNA encoding        tobacco proteinase inhibitor 1), Sumitani et al., Biosci.        Biotech. Biochem. 57:1243 (1993) (nucleotide sequence of        Streptomyces nitrosporeus α-amylase inhibitor) and U.S. Pat. No.        5,494,813 (Hepher and Atkinson, issued Feb. 27, 1996).    -   G. An insect-specific hormone or pheromone such as an        ecdysteroid and juvenile hormone, a variant thereof, a mimetic        based thereon, or an antagonist or agonist thereof. See, for        example, the disclosure by Hammock et al., Nature 344:458        (1990), of baculovirus expression of cloned juvenile hormone        esterase, an inactivator of juvenile hormone.

H. An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest. For example, see thedisclosures of Regan, J. Biol. Chem. 269:9 (1994) (expression cloningyields DNA coding for insect diuretic hormone receptor), and Pratt etal., Biochem. Biophys. Res. Comm. 163:1243 (1989) (an allostatin isidentified in Diploptera puntata). See also U.S. Pat. No. 5,266,317 toTomalski et al., who disclose genes encoding insect-specific, paralyticneurotoxins.

-   -   I. An insect-specific venom produced in nature by a snake, a        wasp, etc. For example, see Pang et al., Gene 116:165 (1992),        fordisclosure of heterologous expression in plants of a gene        coding for a scorpion insectotoxic peptide.    -   J. An enzyme responsible for a hyperaccumulation of a        monterpene, a sesquiterpene, a steroid, hydroxamic acid, a        phenylpropanoid derivative or another non-protein molecule with        insecticidal activity.    -   K. An enzyme involved in the modification, including the        ost-translational modification, of a biologically active        molecule; for example, a glycolytic enzyme, a proteolytic        enzyme, a lipolytic enzyme, a nuclease, a cyclase, a        transaminase, an esterase, a hydrolase, a phosphatase, a kinase,        a phosphorylase, a polymerase, an elastase, a chitinase and a        glucanase, whether natural or synthetic. See PCT application WO        93/02197 in the name of Scott et al., which discloses the        nucleotide sequence of a callase gene. DNA molecules which        contain chitinase-encoding sequences can be obtained, for        example, from the ATCC under Accession Nos. 39637 and 67152. See        also Kramer et al., Insect Biochem. Molec. Biol. 23:691 (1993),        who teach the nucleotide sequence of a cDNA encoding tobacco        hookworm chitinase, and Kawalleck et al., Plant Molec. Biol.        21:673 (1993), who provide the nucleotide sequence of the        parsley ubi4-2 polyubiquitin gene.    -   L. A molecule that stimulates signal transduction. For example,        see the disclosure by Botella et al., Plant Molec. Biol. 24:757        (1994), of nucleotide sequences for mung bean calmodulin cDNA        clones, and Griess et al., Plant Physiol. 104:1467 (1994), who        provide the nucleotide sequence of a maize calmodulin cDNA        clone.    -   M. A hydrophobic moment peptide. See PCT application WO95/16776        (disclosure of peptide derivatives of Tachyplesin which inhibit        fungal plant pathogens) and PCT application WO95/18855 (teaches        synthetic antimicrobial peptides that confer disease        resistance).    -   N. A membrane permease, a channel former or a channel blocker.        For example, see the disclosure of Jaynes et al., Plant Sci        89:43 (1993), of heterologous expression of a cecropin-β, lytic        peptide analog to render transgenic tobacco plants resistant to        Pseudomonas solanacearum.    -   O. A viral-invasive protein or a complex toxin derived        therefrom. For example, the accumulation of viral coat proteins        in transformed plant cells imparts resistance to viral infection        and/or disease development effected by the virus from which the        coat protein gene is derived, as well as by related viruses. See        Beachy et al., Ann. rev. Phytopathol. 28:451 (1990). Coat        protein-mediated resistance has been conferred upon transformed        plants against alfalfa mosaic virus, cucumber mosaic virus,        tobacco streak virus, potato virus X, potato virus Y, tobacco        etch virus, tobacco rattle virus and tobacco mosaic virus. Id.    -   P. An insect-specific antibody or an immunotoxin derived        therefrom. Thus, an antibody targeted to a critical metabolic        function in the insect gut would inactivate an affected enzyme,        killing the insect. Cf. Taylor et al., Abstract #497, Seventh        Int'l Symposium on Molecular Plant-Microbe Interactions        (Edinburgh, Scotland) (1994) (enzymatic inactivation in        transgenic tobacco via production of single-chain antibody        fragments).    -   Q. A virus-specific antibody. See, for example, Tavladoraki et        al., Nature 366:469 (1993), who show that transgenic plants        expressing recombinant antibody genes are protected from virus        attack.    -   R. A developmental-arrestive protein produced in nature by a        pathogen or a parasite. Thus, fungal endo        α-1,4-D-polygalacturonases facilitate fungal colonization and        plant nutrient release by solubilizing plant cell wall        homo-α-1,4-D-galacturonase. See. Lamb et al., Bio/Technology        10:1436 (1992). The cloning and characterization of a gene which        encodes a bean endopolygalacturonase-inhibiting protein is        described by Toubart et al., Plant J. 2:367 (1992).    -   S. A development-arrestive protein produced in nature by a        plant. For example, Logemann et al., Bioi/Technology 10:305        (1992), have shown that transgenic plants expressing the barley        ribosome-inactivting gene have an increased resistance to fungal        disease.    -   2. Genes That Confer Resistance to a Herbicide, For Example:    -   A. A herbicide that inhibits the growing point or meristem, such        as an imidazalinone or a sulfonylurea. Exemplary genes in this        category code for mutant ALS and AHAS enzyme as described, for        example, by Lee et al., EMBO J. 7:1241 (1988), and Miki et al.,        Theor. Appl. Genet. 80:449 (1990), respectively.    -   B. Glyphosate (resistance impaired by mutant        5-enolpyruvl-3-phosphikimate synthase (EPSP) and aroA genes,        respectively) and other phosphono compounds such as glufosinate        (phosphinothricin acetyl transferase, PAT and Streptomyces        hygroscopicus phosphinothricin-acetyl transferase, bar, genes),        and pyridinoxy or phenoxy proprionic acids and cycloshexones        (ACCase inhibitor-encoding genes). See, for example, U.S. Pat.        No. 4,940,835 to Shah, et al., which discloses the nucleotide        sequence of a form of EPSP which can confer glyphosate        resistance. A DNA molecule encoding a mutant aroA gene can be        obtained under ATCC accession number 39256, and the nucleotide        sequence of the mutant gene is disclosed in U.S. Pat. No.        4,769,061 to Comai. European patent application No. 0 333 033 to        Kumada et al., and U.S. Pat. No. 4,975,374 to Goodman et al.,        disclose nucleotide sequences of glutamine synthetase genes        which confer resistance to herbicides such as        L-phosphinothricin. The nucleotide sequence of a        phosphinothricin-acetyl-transferase gene is provided in European        application No. 0 242 246 to Leemans et al., DeGreef et al.,        Bio/Technology 7:61 (1989), describe the production of        transgenic plants that express chimeric bar genes coding for        phosphinothricin acetyl transferase activity. Exemplary of genes        conferring resistance to phenoxy proprionic acids and        cycloshexones, such as sethoxydim and haloxyfop are the Acc1-S1,        Acc1-S2 and Acc1-S3 genes described by Marshall et al., Theor.        Appl. Genet. 83:435 (1992).    -   C. A herbicide that inhibits photosynthesis, such as a triazine        (psbA and gs+genes) and a benzonitrile (nitrilase gene).        Przibila et al., Plant Cell 3:169 (1991), describe the        transformation of Chlamydomonas with plasmids encoding mutant        psbA genes. Nucleotide sequences for nitrilase genes are        disclosed in U.S. Pat. No. 4,810,648 to Stalker, and DNA        molecules containing these genes are available under ATCC        Accession Nos. 53435, 67441, and 67442. Cloning and expression        of DNA coding for a glutathione S-transferase is described by        Hayes et al., Biochem. J. 285:173 (1992).    -   3. Genes That Confer or Contribute to a Value-Added Trait, Such        as:    -   A. Modified fatty acid metabolism, for example, by transforming        a plant with an antisense gene of stearyl-ACP desaturase to        increase stearic acid content of the plant. See Knultzon et al.,        Proc. Natl. Acad. Sci. U.S.A. 89:2624 (1992).    -   B. Decreased phytate content—1) Introduction of a        phytase-encoding gene would enhance breakdown of phytate, adding        more free phosphate to the transformed plant. For example, see        Van Hartingsveldt et al., Gene 127:87 (1993), for a disclosure        of the nucleotide sequence of an Aspergillus niger phytase        gene. 2) A gene could be introduced that reduced phytate        content. In maize, this, for example, could be accomplished, by        cloning and then reintroducing DNA associated with the single        allele which is responsible for maize mutants characterized by        low levels of phytic acid. See Raboy et al., Maydica 35:383        (1990).    -   C. Modified carbohydrate composition effected, for example,        bytransforming plants with a gene coding for an enzyme that        alters the branching pattern of starch. See Shiroza et al., J.        Bacteol. 170:810 (1988) (nucleotide sequence of Streptococcus        mutants fructosyltransferase gene), Steinmetz et al., Mol. Gen.        Genet. 20:220 (1985) (nucleotide sequence of Bacillus subtilis        levansucrase gene), Pen et al., Bio/Technology 10:292 (1992)        (production of transgenic plants that express Bacillus        licheniformis α-amylase), Elliot et al., Plant Molec. Biol.        21:515 (1993) (nucleotide sequences of tomato invertase genes),        Sogaard et al., J. Biol. Chem. 268:22480 (1993) (site-directed        mutagehesis of barley α-amylase gene), and Fisher et al., Plant        Physiol. 102:1045 (1993) (maize endosperm starch branching        enzyme II).

Methods for Soybean Transformation—Numerous methods for planttransformation have been developed, including biological and physical,plant transformation protocols. See, for example, Miki et al.,“Procedures for Introducing Foreign DNA into Plants” in Methods in PlantMolecular Biology and Biotechnology, Glick B. R. and Thompson, J. E.Eds. (CRC Press, Inc., Boca Raton, 1993) pages 67-88. In addition,expression vectors and in vitro culture methods for plant cell or tissuetransformation and regeneration of plants are available. See, forexample, Gruber et al., “Vectors for Plant Transformation” in Methods inPlant Molecular Biology and Biotechnology, Glick B. R. and Thompson, J.E. Eds. (CRC Press, Inc., Boca Raton, 1993) pages 89-119.

A. Agrobacterium-mediated Transformation—One method for introducing anexpression vector into plants is based on the natural transformationsystem of Agrobacterium. See, for example, Horsch et al., Science227:1229 (1985). A. tumefaciens and A. rhizogenesare plant pathogenicsoil bacteria which genetically transform plant cells. The Ti and R1plasmids of A. tumefaciens and A. rhizogenes, respectively, carry genesresponsible for genetic transformation of the plant. See, for example,Kado, C. I., Crit. Rev. Plant Sci. 10:1 (1991). Descriptions ofAgrobacterium vector systems and methods for Agrobacterium-mediated genetransfer are provided by Gruber et al., supra, Miki et al., supra, andMoloney et al., Plant Cell Reports 8:238 (1989). See also, U.S. Pat. No.5,563,055 (Townsend and Thomas), issued Oct. 8, 1996.

B. Direct Gene Transfer— Several methods of plant transformation,collectively referred to as direct gene transfer, have been developed asan alternative to Agrobacterium-mediated transformation. A generallyapplicable method of plant transformation is microprojectile-mediatedtransformation wherein DNA is carried on the surface of microprojectilesmeasuring 1 to 4 μm. The expression vector is introduced into planttissues with a biolistic device that accelerates the microprojectiles tospeeds of 300 to 600 m/s which is sufficient to penetrate plant cellwalls and membranes. Sanford et al., Part. Sci. Technol. 5:27 (1987),Sanford, J. C., Trends Biotech. 6:299 (1988), Klein et al.,Bio/Technology 6:559-563 (1988), Sanford, J. C., Physiol Plant 7:206(1990), Klein et al., Biotechnology 10:268 (1992). See also U.S. Pat.No. 5,015,580 (Christou, et al.), issued May 14, 1991; U.S. Pat. No.5,322,783 (Tomes, et al), issued Jun. 21, 1994.

Another method for physical delivery of DNA to plants is sonication oftarget cells. Zhang et al. Bio/Technology9:996 (1991). Alternatively,liposome or spheroplast fusion have been used to introduce expressionvectors into plants. Deshayes et al., EMBO J., 4:2731 (1985), Christouet al., Proc Natl. Acad. Sci. U.S.A. 84:3962 (1987). Direct uptake ofDNA into protoplasts using CaCl₂ precipitation, polyvinyl alcohol orpoly-L-omithine have also been reported. Hain et al., Mol. Gen. Genet.199:161 (1985) and Draper et al., Plant Cell Physiol. 23:451 (1982).Electroporation of protoplasts and whole cells and tissues have alsobeen described. Donn et al., In Abstracts of VIIth InternationalCongress on Plant Cell and Tissue Culture IAPTC, A2-38, p 53 (1990);D'Halluin et al., Plant Cell 4:1495-1505 (1992) and Spencer et al.,Plant Mol. Biol. 24:51-61 (1994).

Following transformation of soybean target tissues, expression of theabove-described selectable marker genes allows for preferentialselection of transformed cells, tissues and/or plants, usingregeneration and selection methods now well known in the art.

The foregoing methods for transformation would typically be used forproducing a transgenic variety. The transgenic variety could then becrossed, with another (non-transformed or transformed) variety, in orderto produce a new transgenic variety. Alternatively, a genetic traitwhich has been engineered into a particular soybean line using theforegoing transformation techniques could be moved into another lineusing traditional backcrossing techniques that are well known in theplant breeding arts. For example, a backcrossing approach could be usedto move an engineered trait from a public, non-elite variety into anelite variety, or from a variety containing a foreign gene in its genomeinto a variety or varieties which do not contain that gene. As usedherein, “crossing” can refer to a simple X by Y cross, or the process ofbackcrossing, depending on the context.

Single Gene Conversions—When the term soybean plant is used in thecontext of the present invention, this also includes any single geneconversions of that variety. The term single gene converted plant asused herein refers to those soybean plants which are developed by aplant breeding technique called backcrossing wherein essentially all ofthe desired morphological and physiological characteristics of a varietyare recovered in addition to the single gene transferred into thevariety via the backcrossing technique. Backcrossing methods can be usedwith the present invention to improve or introduce a characteristic intothe variety. The term backcrossing as used herein refers to the repeatedcrossing of a hybrid progeny back to the recurrent parent, i.e.,crossing back 1, 2, 3, 4, 5, 6, 7, 8 or more times to the recurrentparent. The parental soybean plant which contributes the gene for thedesired characteristic is termed the nonrecurrent or donor parent. Thisterminology refers to the fact that the nonrecurrent parent is used onetime in the backcross protocol and therefore does not recur. Theparental soybean plant to which the gene or genes from the nonrecurrentparent are transferred is known as the recurrent parent as it is usedfor several rounds in the backcrossing protocol (Poehlman & Sleper,1994; Fehr, 1987). In a typical backcross protocol, the original varietyof interest (recurrent parent) is crossed to a second variety(nonrecurrent parent) that carries the single gene of interest to betransferred. The resulting progeny from this cross are then crossedagain to the recurrent parent and the process is repeated until asoybean plant is obtained wherein essentially all of the desiredmorphological and physiological characteristics of the recurrent parentare recovered in the converted plant, in addition to the singletransferred gene from the nonrecurrent parent.

The selection of a suitable recurrent parent is an important step for asuccessful backcrossing procedure. The goal of a backcross protocol isto alter or substitute a single trait or characteristic in the originalvariety. To accomplish this, a single gene of the recurrent variety ismodified or substituted with the desired gene from the nonrecurrentparent, while retaining essentially all of the rest of the desiredgenetic, and therefore the desired physiological and morphological,constitution of the original variety. The choice of the particularnonrecurrent parent will depend on the purpose of the backcross, one ofthe major purposes is to add some commercially desirable, agronomicallyimportant trait to the plant. The exact backcrossing protocol willdepend on the characteristic or trait being altered to determine anappropriate testing protocol. Although backcrossing methods aresimplified when the characteristic being transferred is a dominantallele, a recessive allele may also be transferred. In this instance itmay be necessary to introduce a test of the progeny to determine if thedesired characteristic has been successfully transferred.

Many single gene traits have been identified that are not regularlyselected for in the development of a new variety but that can beimproved by backcrossing techniques. Single gene traits may or may notbe transgenic, examples of these traits include but are not limited to,male sterility, waxy starch, herbicide resistance, resistance forbacterial, fungal, or viral disease, insect resistance, male fertility,enhanced nutritional quality, industrial usage, yield stability andyield enhancement. These genes are generally inherited through thenucleus. Several of these single gene traits are described in U.S. Pat.Nos. 5,959,185, 5,973,234 and 5,977,445, the disclosures of which arespecifically hereby incorporated by reference.

Further reproduction of the variety can occur by tissue culture andregeneration. Tissue culture of various tissues of soybeans andregeneration of plants therefrom is well known and widely published. Forexample, reference may be had to Komatsuda, T. et al., “Genotype XSucrose Interactions for Somatic Embryogenesis in Soybean,” Crop Sci.31:333-337 (1991); Stephens, P. A., et al., “Agronomic Evaluation ofTissue-Culture-Derived Soybean Plants,” Theor. Appl. Genet. (1991)82:633-635; Komatsuda, T. et al., “Maturation and Germination of SomaticEmbryos as Affected by Sucrose and Plant Growth Regulators in SoybeansGlycine gracilis Skvortz and Glycine max (L.) Merr.” Plant Cell, Tissueand Organ Culture, 28:103-113 (1992); Dhir, S. et al., “Regeneration ofFertile Plants from Protoplasts of Soybean (Glycine max L. Merr.);Genotypic Differences in Culture Response,” Plant Cell Reports (1992)11:285-289; Pandey, P. et al., “Plant Regeneration from Leaf andHypocotyl Explants of Glycine-wightii (W. and A.) VERDC. var.longicauda,” Japan J. Breed. 42:1-5 (1992); and Shetty, K., et al.,“Stimulation of In Vitro Shoot Organogenesis in Glycine max (Merrill.)by Allantoin and Amides,” Plant Science 81:245-251 (1992); as well asU.S. Pat. No. 5,024,944 issued Jun. 18, 1991 to Collins et al., and U.S.Pat. No. 5,008,200 issued Apr. 16, 1991 to Ranch et al. Thus, anotheraspect of this invention is to provide cells which upon growth anddifferentiation produce soybean plants having the physiological andmorphological characteristics of soybean variety S030159.

As used herein, the term “tissue culture” indicates a compositioncomprising isolated cells of the same or a different type or acollection of such cells organized into parts of a plant. Exemplarytypes of tissue cultures are protoplasts, calli, plant clumps, and plantcells that can generate tissue culture that are intact in plants orparts of plants, such as embryos, pollen, flowers, seeds, pods, leaves,stems, roots, root tips, anthers, and the like. Means for preparing andmaintaining plant tissue culture are well known in the art. By way ofexample, a tissue culture comprising organs has been used to produceregenerated plants. U.S. Pat. Nos. 5,959,185; 5,973,234 and 5,977,445,described certain techniques, the disclosures of which are incorporatedherein by reference.

This invention also is directed to methods for producing a soybean plantby crossing a first parent soybean plant with a second parent soybeanplant wherein the first or second parent soybean plant is a soybeanplant of the variety S030159. Further, both first and second parentsoybean plants can come from the soybean variety S030159. Thus, any suchmethods using the soybean variety S030159 are part of this invention:selfing, backcrosses, hybrid production, crosses to populations, and thelike. All plants produced using soybean variety S030159 as a parent arewithin the scope of this invention, including those developed fromvarieties derived from soybean variety S030159. Advantageously, thesoybean variety could be used in crosses with other, different, soybeanplants to produce first generation (F₁) soybean hybrid seeds and plantswith superior characteristics. The variety of the invention can also beused for transformation where exogenous genes are introduced andexpressed by the variety of the invention. Genetic variants createdeitherthrough traditional breeding methods using variety S030159 orthrough transformation of S030159 by any of a number of protocols knownto those of skill in the art are intended to be within the scope of thisinvention.

As used herein, the term plant includes plant cells, plant protoplasts,plant cell tissue cultures from which soybean plants can be regenerated,plant calli, plant clumps, and plant cells that are intact in plants orparts of plants, such as embryos, pollen, ovules, flowers, pods, leaves,roots, root tips, anthers, and the like.

Industrial Uses

The seed of soybean variety S030159, the plant produced from the seed,the hybrid soybean plant produced from the crossing of the variety withany other soybean plant, hybrid seed, and various parts of the hybridsoybean plant can be utilized for human food, livestock feed, and as araw material in industry.

The soybean is the world's leading source of vegetable oil and proteinmeal. The oil extracted from soybeans is used for cooking oil,margarine, and salad dressings. Soybean oil is composed of saturated,monounsaturated and polyunsaturated fatty acids. It has a typicalcomposition of 11% palmitic, 4% stearic, 25% oleic, 50% linoleic and 9%linolenic fatty acid content (“Economic Implications of Modified SoybeanTraits Summary Report”, Iowa Soybean Promotion Board and AmericanSoybean Association Special Report 92S, May 1990). Changes in fatty acidcomposition for improved oxidative stability and nutrition areconstantly sought after. Industrial uses of soybean oil which issubjected to further processing include ingredients for paints,plastics, fibers, detergents, cosmetics and lubricants. Soybean oil maybe split, inter-esterified, sulfurized, epoxidized, polymerized,ethoxylated, or cleaved. Designing and producing soybean oil derivativeswith improved functionality, oliochemistry, is a rapidly growing field.The typical mixture of triglycerides is usually split and separated intopure fatty acids, which are then combined with petroleum-derivedalcohols or acids, nitrogen, sulfonates, chlorine, or with fattyalcohols derived from fats and oils.

Soybean is also used as a food source for both animals and humans.Soybean is widely used as a source of protein for animal feeds forpoultry, swine and cattle. During processing of whole soybeans, thefibrous hull is removed and the oil is extracted. The remaining soybeanmeal is a combination of carbohydrates and approximately 50% protein.

For human consumption soybean meal is made into soybean flour which isprocessed to protein concentrates used for meat extenders or specialtypet foods. Production of edible protein ingredients from soybean offersa healthy, less expensive replacement for animal protein in meats aswell as dairy-type products.

Tables

In Table 2 that follows, the traits and characteristics of soybeancultivar S030159 are compared to several competing varieties ofcommercial soybeans of similar maturity. In the tables, column 1 showsthe comparison number; column 2 is the year of the test; columns 3 and 4give the number of locations and number of observations, respectively.Column 5 indicates the genotype and column 6 shows the mean yield.Column 7 presents the t value and columns 8 and 9 present the critical tvalues at the 0.05% and 0.01% levels of significance, respectively.

As shown in Table 2, soybean cultivar S030159 yields higher than 15commercial varieties with the increase over all comparisons beingsignificant at the 0.01 level of probability except CSR2533 and DKB26-52where the increase is significant at the 0.05 level and AG2403 where theincrease is not significant.

TABLE 2 PAIRED COMPARISONS Mean Critical Critical Comp # Year # of Loc.# of Obs. Genotype Yield t Value t @ .05 t @ .01 1 2003 16 48 S03015943.6 3.51** 1.68 2.41 CSR2104-03 40.5 2 2003 16 48 S030159 43.6 4.52**1.68 2.41 CSR2312N 40.0 3 2003 16 48 S030159 43.6 3.62** 1.68 2.41P92B38 39.8 4 2003 16 48 S030159 43.6 2.75** 1.68 2.41 CSR2320 40.7 52003 16 48 S030159 43.6 4.40** 1.68 2.41 CSR2423 38.9 6 2003 16 48S030159 43.6 1.50 1.68 2.41 AG2403 41.9 7 2003 16 48 S030159 43.6 4.98**1.68 2.41 AG2405 38.7 8 2003 16 48 S030159 43.6 4.59** 1.68 2.41MBS50462NRR 39.5 9 2003 16 47 S030159 43.7 3.60** 1.68 2.41 NKS24-K439.8 10 2003 16 47 S030159 437 5.53** 1.68 2.41 P92B47 38.3 11 2003 1648 S030159 43.6 4.86** 1.68 2.41 CSR2530 39.1 12 2003 16 48 S030159 43.61.83* 1.65 2.41 CSR2533 41.4 13 2003 16 48 S030159 43.6 2.90** 1.68 2.41JG925012 40.9 14 2003 16 48 S030159 43.6 3.63** 1.68 2.41 CSR2520 40.215 2003 16 48 S030159 43.6 2.13* 1.68 2.41 DKB26-52 41.5 *Significant at.05 level of probability **Significant at .01 level of probability

Deposit Information

A deposit of the Stine Seed Farm, Inc. and Monsanto Technology LLCproprietary soybean cultivar S030159 disclosed above and recited in theappended claims has been made with the American Type Culture Collection(ATCC), 10801 University Boulevard, Manassas, Va. 20110. The date ofdeposit was Dec. 28, 2004. The deposit of 2,500 seeds was taken from thesame deposit maintained by Stine Seed Farm, Inc. since prior to thefiling date of this application. All restrictions upon the deposit havebeen removed, and the deposit is intended to meet all of therequirements of 37 C.F.R. §1.801-1.809. The ATCC accession number isPTA-6485. The deposit will be maintained in the depository for a periodof 30 years, or 5 years after the last request , or for the effectivelife of the patent, whichever is longer, and will be replaced asnecessary during that period.

Although the foregoing invention has been described in some detail byway of illustration and example for purposes of clarity andunderstanding. However, it will be obvious that certain changes andmodifications such as single gene modifications and mutations,somoclonal variants, variant individuals selected from large populationsof the plants of the instant variety and the like may be practicedwithin the scope of the invention, as limited only by the scope of theappended claims.

1. Seed of soybean line designated S030159, representative seed of saidline having been deposited under ATCC Accession No. PTA-6485.
 2. Asoybean plant, or a part thereof, produced by growing the seed ofclaim
 1. 3. A tissue culture of regenerable cells produced from theplant of claim
 2. 4. Protoplasts produced from the tissue culture ofclaim
 3. 5. The tissue culture of claim 3, wherein cells of the tissueculture are produced from a tissue selected from the group consisting ofleaf, pollen, embryo, root, root tip, anther, pistil, flower, seed, pod,and stem.
 6. A soybean plant regenerated from the tissue culture ofclaim 3, said plant having all the morphological and physiologicalcharacteristics of line S030159, representative seed of said line havingbeen deposited under ATCC Accession No. PTA-6485.
 7. A method forproducing an F1 hybrid soybean seed, comprising crossing the plant ofclaim 2 with a different soybean plant and harvesting the resultant F1hybrid soybean seed.
 8. A method for producing a male sterile soybeanplant comprising transforming the soybean plant of claim 2 with anucleic acid molecule that confers male sterility.
 9. A male sterilesoybean plant produced by the method of claim
 8. 10. A method ofproducing an herbicide resistant soybean plant comprising transformingthe soybean plant of claim 2 with a transgene that confers herbicideresistance.
 11. An herbicide resistant soybean plant produced by themethod of claim
 10. 12. The soybean plant of claim 11, wherein thetransgene confers resistance to an herbicide selected from the groupconsisting of imidazolinone, sulfonylurea, glyphosate, glufosinate,L-phosphinothricin, triazine and benzonitrile.
 13. A method of producingan insect resistant soybean plant comprising transforming the soybeanplant of claim 2 with a transgene that confers insect resistance.
 14. Aninsect resistant soybean plant produced by the method of claim
 13. 15.The soybean plant of claim 14, wherein the transgene encodes a Bacillusthuringiensis endotoxin.
 16. A method of producing a disease resistantsoybean plant comprising transforming the soybean plant of claim 2 witha transgene that confers disease resistance.
 17. A disease resistantsoybean plant produced by the method of claim
 16. 18. A soybean plant,or part thereof, having all the physiological and morphologicalcharacteristics of the line S030159, representative seed of said linehaving been deposited under ATCC Accession No. PTA-6485.
 19. A method ofproducing a soybean plant with modified fatty acid or carbohydratemetabolism wherein the method comprises transforming the soybean plantof claim 2 with a transgene encoding a protein selected from the groupconsisting of fructosyltransferase, levansucrase, α-amylase, invertaseand starch branching enzyme or encoding an antisense of steayl-ACPdesaturase.
 20. A soybean plant having modified fatty acid orcarbohydrate metabolism produced by the method of claim 19.